Project Summary/Abstract This is an SBIR Phase I proposal to test the feasibility of using a novel small molecule drug for the treatment of a pediatric rare cancer, retinoblastoma (Rb). Rb is currently treated using drug combinations in concert with non-drug treatments. Rb occurs in most cases before age 3. Current cytotoxic drug treatments produce high toxicities to childhood patients. Moreover, Rb tends to metastasize (break off and seeding), creating an additional challenge for management of this disease. Conclusively, a drug with low toxicity and high efficacy against primary and secondary cancers would help to avoid the disadvantages of current chemotherapies and potentially decrease mortality induced by the disease. In this project, we will use human Rb tumor models developed from both primary and metastatic disease sites to test a novel small molecule drug for Rb treatment. Previous studies reveal that Rb has an intact p53 pathway, but this pathway is constitutively inhibited through increased levels of the oncogenic proteins Mdm4/MdmX and survivin. Furthermore, genomic and epigenetic analyses of Rb found that the proto-oncoprotein SYK is usually not expressed in normal human retina, but is activated in Rb, and in turn decrease apoptosis via induction of Mcl-1 expression. We have developed a small molecule FL118 that effectively inhibits the expression of Mcl-1, survivin and MdmX, as well as XIAP, and cIAP2 in colon cancer cells. Furthermore, FL118 rapidly induces p53 protein accumulation via inducing the degradation of MdmX in colon cancer cells. These features of FL118 suggest a potential for the treatment of Rb. However, in consideration of the fact that Rb is a pediatric cancer, we wondered whether we could find a more Rb-targeted FL118 analogue with even lower toxicity and higher efficacy. To this end, we synthesized a focused library of 35 FL118 analogues and tested all of them in vivo for their efficacy and initial toxicity with the Rb cell line HT-3-established xenograft tumors in mice. We identified three FL118 analogues (FL7N-1, FL7N-2 and FL7N-3) that showed high efficacy to inhibit HT-3 xenograft tumor growth without obvious toxicity to animals. FL7N-1 effectively inhibits survivin, MdmX and Mcl-1 and induces p53 in Rb cells. To achieve our Specific Aim of testing the feasibility of FL7N-1 for Rb treatment, we will carry out the following three Tasks. Task 1: Determine FL7N-1 toxicity and maximum tolerated dose (MTD) using animal models. Task 2: Determine FL7N-1 dose response and therapeutic index (TI) using Rb-bearing animals. Task 3: Determine FL7N-1 pharmacokinetics (PK) profiles in Rb-bearing mice. Test of Feasibility: We must observe that the FL7N-1 MTD in immuno-competent animals is equal to or higher than in immuno-compromised mice. The FL7N-1 most effective dose (MED) must be lower than its MTD. Using our conservative TI definition [MTD divided by MED], the TI for FL7N-1 must be >1. In SBIR Phase II, we will carry out additional preclinical studies to define the best therapeutic schedule with dose range, route of administration, and other IND-enabling studies for filing an FL7N-1 IND for clinical trials.